Department of Chemical & Biomolecular Engineering, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, IL 61801, USA.
Biosens Bioelectron. 2013 Nov 15;49:118-25. doi: 10.1016/j.bios.2013.04.046. Epub 2013 May 9.
Effective treatment of clinical infections is critically dependent on the ability to rapidly screen patient samples to identify antibiograms of infecting pathogens. Existing methods for antibiotic susceptibility testing suffer from several disadvantages, including long turnaround times, excess sample and reagent consumption, poor detection sensitivity, and limited combinatorial capabilities. Unfortunately, these factors preclude the timely administration of appropriate antibiotics, complicating management of infections and exacerbating the development of antibiotic resistance. Here, we seek to address these issues by developing a microfluidic platform that relies on fluorescence detection of bacteria that express green fluorescent protein for highly sensitive and rapid antibiotic susceptibility testing. This platform possesses several advantages compared to conventional methods: (1) analysis of antibiotic action in two to four hours, (2) enhanced detection sensitivity (≈ 1 cell), (3) minimal consumption of cell samples and antibiotic reagents (<6 µL), and (4) improved portability through the implementation of normally closed valves. We employed this platform to quantify the effects of four antibiotics (ampicillin, cefalexin, chloramphenicol, tetracycline) and their combinations on Escherichia coli. Within four hours, the susceptibility of bacteria to antibiotics can be determined by detecting variations in maxima of local fluorescence intensity over time. As expected, cell density is a major determinant of antibiotic efficacy. Our results also revealed that combinations of three or more antibiotics are not necessarily better for eradicating pathogens compared to pairs of antibiotics. Overall, this microfluidic based biosensor technology has the potential to provide rapid and precise guidance in clinical therapies by identifying the antibiograms of pathogens.
临床感染的有效治疗严重依赖于快速筛选患者样本以确定感染病原体的抗生素药敏谱的能力。现有的抗生素药敏检测方法存在几个缺点，包括周转时间长、样本和试剂消耗过多、检测灵敏度差以及组合能力有限。不幸的是，这些因素妨碍了及时给予适当的抗生素，使感染的管理变得复杂，并加剧了抗生素耐药性的发展。在这里，我们通过开发一种依赖于表达绿色荧光蛋白的细菌的荧光检测来进行高度敏感和快速抗生素药敏检测的微流控平台来解决这些问题。与传统方法相比，该平台具有以下几个优势：（1）在两到四个小时内分析抗生素的作用，（2）增强检测灵敏度（≈1 个细胞），（3）最小化细胞样本和抗生素试剂的消耗（<6 µL），以及（4）通过实施常闭阀提高便携性。我们使用该平台来量化四种抗生素（氨苄青霉素、头孢氨苄、氯霉素、四环素）及其组合对大肠杆菌的影响。在四个小时内，可以通过检测随时间变化的局部荧光强度最大值来确定细菌对抗生素的敏感性。正如预期的那样，细胞密度是抗生素疗效的主要决定因素。我们的结果还表明，与两种抗生素相比，三种或更多种抗生素的组合不一定更有利于消灭病原体。总的来说，这种基于微流控的生物传感器技术有可能通过确定病原体的抗生素药敏谱为临床治疗提供快速和精确的指导。
